Tetrahydrobiopterin (BH4) biosynthesis has been shown to proceed through tetrahydro intermediates. The first tetrahydro intermediate has been shown to be 6-pyruvoyl-tetrahydropterin. The enzyme catalyzing the formation of this intermediate, 6-pyruvoyl-tetrahydropterin synthase, has been purified from brain and liver. Sepiapterin reductase catalyzes the last reaction in BH4 biosynthesis.